CX3CL1 Induces Vertebral Microvascular Barrier Dysfunction via the Src/P115-RhoGEF/ROCK Signaling Pathway.

Lei Yi,Yun Liang,Houlei Wang,Quanming Zhao,Jian Dong

doi:10.3389/fncel.2020.00096

Abstract

Trans-endothelial migration (TEM) of cancer cells is a critical step in metastasis. Micro-vascular barrier disruptions of distant organs play important roles in tumor cells TEM. The spine is a preferred site for multiple cancer cell metastases. Our previous study found that vertebral spongy bone was rich in CX3CL1 and that CX3CL1 can attract fractalkine receptor-expressing tumor cells to the spine. In the present study, we determined whether CX3CL1 was involved in vertebral micro-vascular barrier disruption and promoted tumor cell TEM after circulating tumor cells were arrested in the vertebral micro-vasculature. We examined the role of CX3CL1 in the barrier function of vertebral micro-vascular endothelial cells (VMECs) and explored the molecular mechanisms of CX3CL1-induced VMEC barrier disruption. Our results demonstrated that CX3CL1 led to F-actin formation and ZO-1 disruption in VMECs and induced the vertebral micro-vascular barrier disruption. Importantly, we found that the activation of the Src/P115-RhoGEF/ROCK signaling pathway plays an important role in CX3CL1-induced VMEC stress fiber formation, ZO-1 disruption and then vertebral micro-vascular barrier hyper-permeability. Inhibiting Src/P115-RhoGEF/ROCK signaling in VMECs effectively blocked CX3CL1-induced vertebral vascular endothelial dysfunction and subsequent tumor cell TEM. The results of this study and our previous study indicate that in addition to its chemotaxis, CX3CL1 plays a critical role in regulating vertebral micro-vascular barrier function and tumor cell TEM. CX3CL1 induced VMECs stress fiber formation, ZO-1 disruption and then vascular endothelial hyperpermeability via activation of the Src/P115-RhoGEF/ROCK signaling pathway. The inhibition of the Src/P115-RhoGEF/ROCK signaling pathway in VMECs effectively blocked tumor cells TEMs in vertebral spongy bone and maybe a potential therapeutic strategy for spine metastases in the future.

Highlights

Metastasis is a fatal step in the progression of many malignancies
The measurements revealed that CX3CL1 induced VMEC barrier disruption, trans-endothelial electrical resistance (TER) decreased in a dose- and timedependent manner and peak EC barrier disruption appeared at 6 h after stimulation with 1 μg/ml CX3CL1 (Figures 2A,B)
We found that only incubating VMECs with CX3CL1 increased the trans-endothelial migration (TEM) of tumor cells but that pre-inhibition of Src/P115-RhoGEF/Rho-associated protein kinase (ROCK) signaling in the VMECs significantly reversed CX3CL1-induced VMECs barrier hyperpermeability and the high rate of tumor cell TEM

Summary

Introduction

Metastasis is a fatal step in the progression of many malignancies. Vertebral cancellous bones, as a preferential district for bone metastases, are highly efficient at arresting circulating cancer cells from low-velocity bloodstream. CX3CL1 Induces VMECs Barrier Disruption functions as an adhesion molecule for arresting circulating tumor cells in spine microvasculature (Ostuni et al, 2014; Zhou et al, 2016). The role of CX3CL1 in tumor cell trans-endothelial migration (TEM) in the spine remains unknown. After circulating cancer cells arrested in the microvasculature in the spine, the extravasation of circulating tumor cells into the vertebral spongy bone is a critical step for metastasis. In tumor cell TEM, microvascular endothelial barrier dysfunction and subsequent micro-vascular hyper-permeability promote metastasis formation. In this study, we hypothesized that CX3CL1 induced the formation of stress fiber, the ZO-1 disruption in VMECs and promoted VMECs barrier disruption, which further mediated vertebral micro-vascular hyper-permeability and increased tumor cell TEM in the spine

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